The present invention relates to a novel flavone derivative represented by the following formula (1): 
in which
R1, R3 and R4 each independently represent hydrogen, halogen, hydroxy, alkyl, lower alkoxy, amino or nitro,
R2 represents hydrogen, halogen, hydroxy, lower alkyl, lower alkoxy, amino, nitro or 
xe2x80x83wherein
A represents amino which may be optionally substituted with alkyl, cycloalkyl, aralkyl, acyl, or aryl which is optionally substituted with one or two substituents selected from the group consisting of halogen, cyano, nitro and amino; or 1,2,3,4-tetrahydroisoquinoline which may be optionally substituted with halogenoalkylcarbonyl; or alkyl, aryl, aralkyl or heteroaryl each of which may be optionally substituted with one or two substituents selected from the group consisting of alkyl, halogenoalkyl, halogen, dialkylamino, phenyl, nitro, amino, isooxazole, pyridine, carboxy, morpholine, methylpiperazine and cyano,
Y represents SO2 or CO,
B represents hydrogen or alkyl,
R5 represents hydrogen or hydroxy, and
R6 and R7 are substituted at o-, m- or p-position from each other and each independently represents hydrogen, hydroxy, halogen or lower alkoxy or together represent lower alkylenedioxy,
pharmaceutically acceptable salt, hydrate, solvate and isomer thereof which is useful as an inhibitor for Cyclin Dependent Kinase (hereinafter, referred to as xe2x80x9cCDKxe2x80x9d),
In addition, the present invention relates to a process for preparing the flavone derivative of formula (1) and also relates to an anti-cancer agent or an agent for treating neurodegenerative disease characterized by comprising the compound of formula (1) as an active ingredient.
Researches on cell division process in molecular level have been extensively performed from the late 1980""s through study of division of frog oocytes, analysis several yeast cell growth or characterization of induced mutants by radiation and study of the tumor suppressor Rb. In the 1990""s, it is discovered that small molecular cell growth regulator controls cell division process (i.e. growth, differentiation, cytogenesis, aging and apoptosis etc.) through its own regulatory function. These results were very useful for more precise understanding of the pathology of several diseases.
A representative example is cancer. In transformation process from normal cells to cancer cells, it was frequently observed that cell growth regulator loses its own function. That is to say, in cancer cells, the cell growth regulator shows an abnormal activity, which is deeply associated with invasion/metastasis which is crucial in the cancerpathology. Particularly, cell cycle deregulation is recognized to be a direct cause of cancer since cancer occurs in experimental animal when overexpression or knock-out of cell growth regulator is induced by using tranformed animal.
The cell growth is under positive or negative regulation in the same manner as other biological regulations. The major pathway of cell cycle regulation known up to now is based on CDK activity and as a result of studies on many cancer cells and carcinogenesis mechanisms, it was confirmed that problems of positive or negative regulation on CDK activity result in carcinogenesis in many cases. That is, cancer may occur when positive or negative regulation and timely regulation which is important for cell growth regulation are disrupted.
The representative CDKs of mammals are CDK4 (Cyclin dependent kinase 4) which shows its activity in mid-G1 phase of cell cycle, CDK2 which shows its activity in mid-1 and S phases, CDC2 (CDK1) which shows its activity in G2-M phase, and so on. It is known that CDK4 and CDK2 activities are regulated by check point of G1-S cell cycle and CDC2 activity by check point of G2-M. In many cancer cells, abnormalities appear in the regulatory mechanism of CDK4, CDK2 and CDC2 (CDK 1) and in fact, it was confirmed that induced abnormalities cause cancer in the transformed animal. Therefore, CDK4, CDK2 and CDC2 (CDK1) among several kinds of CDKs are suitable as a target of anti-cancer agents.
The results of studies on relation between these CDKs and carcinogtenesis will be explained in more detail in the following.
The relation between the abnormal regulation of CDK4 activity and carcinogenesis is observed in several cancer tissues. The deletion of p16 and p15 genes in several kinds of cancer is reported and particularly, overexpression of cyclin D1 is observed, which has close relation with the fact that breast cancer has a metastatic proper and which suggests that malignant phenotype may be expressed when CDK4 activity is deregulated.
Furthermore, it was reported that p16 knocked-out mouse has such a high carcinogenesis rate as p53 knocked-out mouse, which suggests that malfunction of p16 on CDK4 regulation is a cause of carcinogenesis. It gives the possibility that p16 plays a role in the downstream in NIH 3T3 cell with overexpressed ras or src. Reversely it was observed that modified phenotype wherein p16 or p21 is transformed with ras is repaired into wild phenotype. From these experimental results, deregulation of CDK4 activity may be a cause of carcinogenesis and play a role in maintenance of phenotype of cancer cell. Therefore, CDK4 inhibitors may have anti-cancer effects.
It was reported that overexpression of cyclin E is observed in some breast cancers, deeply associated with metastasis of breast cancer, inhibits cell apoptosis under low serum condition and induces anchorage independent growth, and that hyperproliferation (neoplasia) of mammary epithelial cells is observed in transformed animal with overexpressed CDK2 by MMTV promoter, which suggests that CDK2 activity is related with the progress or maintenance of cell transformation and CDK2 inhibitors may also have anti-cancer effects.
It is recently discovered that CDK5 among these CDKs may cause neurodegenerative diseases by phosphorylation of tau protein of the brain. Therefore, CDK5 inhibitor may be useful as an agent for treating neurodegenerative diseases (e.g. Alzheimer""s disease). CDK2 inhibitors may have inhibitory effects on CDK5 in that CDK2 and CDK5 are homologous in the same family (Ref.: John Leu et al., xe2x80x9cNeuronal CDC2-like kinasexe2x80x9d, TIBS, January 1995, pp33xcx9c37).
Furthermore, it is discovered that CDC2 (CDK1), CDK3, CDK6 and CDK7 play an important role in each phase of cell division. These are classified into CDKs family. In addition, to cyclin D1 and E, cyclin A, B, C, D2, D3, D4, F and G are also calssified into the same family.
On the basis of the above-mentioned research, efficient inhibitors of these CDKs may be useful as anti-cancer or anti-neurodegenerative agents. Therefore, recently, these inhibitors have been developed.
As effective CDK inhibitors developed hitherto, there exists Flavopiridol, compound of the formula (2) 
which is under clincal trials [Ref.: EP 0,241,003 and 0,336,061]. In addition, a purine derivative of the formula (3) 
has been recently developed [Ref: WO 97/16447].
However, the CDK inhibitors developed up to now could not have satisfactory effects.
So, the present inventors have made widespread and concentrative researches on CDK inhibitors and as a result, found that the above flavone derivative of formula (1) which has a quite different structure from any other known CDK inhibitors inhibits CDKs effectively and finally. complete the present invention.
The object of the present invention is to provide a novel flavone derivative of formula (1), pharmaceutically acceptable salt, hydrate, solvate and isomer thereof having an inhibitory activity for CDK.
Another object of the present invention is to provide a process for preparing the compound of formula (1). Still another object of the present invention is to provide an anti-cancer agent and an agent for treating neurodegenerative disease each of which is characterized by comprising as an active ingredient the compound of formula (1) with a pharmaceutically acceptable carrier.
In this specification, CDKs includes all of CDK2, CDK4, CDC2 (CDK1), CDK3, CDK5, CDK6, CDK7 etc., and cyclin includes cyclin D1, E, A, B, C, D2, D3, D4, F, and G.
The present invention is explained in detail hereinafter.
The present invention relates to a novel flavone derivative represented by the following formula (1): 
in which
R1, R3 and R4 each independently represent hydrogen, halogen. hydroxy, alkyl, lower alkoxy, amino or nitro.
R2 represents hydrogen, halogen, hydroxy, lower alkyl, lower alkoxy, amino, nitro or 
xe2x80x83wherein
A represents amino which may be optionally substituted with alkyl, cycloalkyl, aralkyl, acyl, or aryl which is optionally substituted with one or two substituents selected from the group consisting of halogen, cyano, nitro and amino; or 1,2,3,4-tetrahydroisoquinoline which may be optionally substituted with halogenoalkylcarbonyl; or alkyl, aryl, aralkyl or heteroaryl each of which may be optionally substituted with one or two substituents selected from the group consisting of alkyl, halogenoalkyl, halogen, dialkylamino, phenyl, nitro, amino, isooxazole, pyridine, carboxy, morpholine, methylpiperazine and cyano,
Y represents SO2 or CO,
B represents hydrogen or alkyl,
R5 represents hydrogen or hydroxy, and
R6 and R7 are substituted at o-, m- or p-position from each other and each independently represents hydrogen, hydroxy, halogen or lower alkoxy or together represent lower alkylenedioxy,
pharmaceutically acceptable salt, hydrate, solvate and isomer thereof which has efficacies on treating cancer or neurodegenerative disease through a mechanism of inhibiting CDKs activity,
In addition, the present invention relates to a process for preparing the flavone derivative of formula (1) and also relates to an anti-cancer agent or an agent for treating neurodegenerative disease each of which is characterized by comprising the compound of formula (1) as an active ingredient.
Among the compound of formula (1) according to the present invention, the preferred compounds include those wherein
(1) R1, R2, R3 and R4 each independently represent hydrogen, halogen. hydroxy, lower alkoxy or nitro (provided that two or more of these four substituents are hydrogen),
R5 represents hydrogen or hydroxy, and
R6 and R7 are substituted at o-, m- or p-position from each other and each independently represents hydrogen, hydroxy, halogen or lower alkoxy or together represent lower alkylenedioxy, or
(2) R1 represents hydrogen, hydroxy, lower alkoxy or amino,
R2 and R3 each independently represent hydrogen, halogen, hydroxy, lower alkyl, lower alkoxy or amino,
R4 represents hydrogen, halogen, hydroxy or lower alkoxy,
R5 represents hydroxy, and
R6 and R7 represent 3-hydroxy and 4-hydroxy respectively, or
(3) R1, R3 and R4 each independently represent hydrogen, halogen, hydroxy, alkyl or amino (provided that two or more of these three substituents are hydrogen),
R2 represents 
xe2x80x83wherein
A represents amino which may be optionally substituted with alkyl, cycloalkyl, aralkyl, acyl, or amyl which is optionally substituted with one or two substituents selected from the group consisting of halogen, cyano, nitro and amino; or 1,2,3,4-tetrahydroisoquinoline which may be optionally substituted with halogenoalkylcarbonyl; or alkyl, aryl, aralkyl or heteroaryl each of which may be optionally substituted with one or two substituents selected from the group consisting of alkyl, halogenoalkyl, halogen, dialkylamino, phenyl, nitro, amino, isooxazole, pyridine, carboxy, morpholine, methylpiperazine and cyano,
Y represents SO2 or CO,
B represents hydrogen or alkyl,
R5 represents hydroxy, and
R6 and R7 represent 3-hydroxy and 4-hydroxy respectively.
More preferred compounds include those wherein
(1) R1, R2, R3 and R4 each independently represent hydrogen, chloro, hydroxy, methoxy or nitro(provided that two or more of these four substituents are hydrogen),
R5 represents hydrogen or hydroxy, and
R6 and R7 are substituted at o-, m- or p-position from each other and each independently represents hydrogen, hydroxy, chloro or methoxy or together represent methylenedioxy, or
(2) R1 represents hydrogen, hydroxy or lower alkoxy,
R2 represents hydrogen, halogen, hydroxy, lower alkyl, lower alkoxy or amino,
R3 represents hydrogen, hydroxy, lower alkyl, lower alkoxy or amino,
R4 represents hydrogen, halogen or hydroxy,
R5 represents hydroxy, and
R6, and R7 represent 3-hydroxy and 4-hydroxy respectively, or
(3) R1, R3 and R4 each represent hydrogen,
R2 represents 
xe2x80x83wherein
A represents amino which may be optionally substituted with cycloalkyl; or 1,2,3,4-tetrahydroisoquinoline which may be optionally substituted with halogenoalkylcarbonyl; or alkyl, aryl, aralkyl or heteroaryl each of which may be optionally substituted with one or two substituents selected from the group consisting of alkyl, halogenoalkyl, halogen, dialkylamino, phenyl. nitro, amino isooxazole, pyridine carboxy, morpholine and methylpiperazine,
Y represents SO2,
B represents hydrogen,
R5 represents hydroxy, and
R6 and R7 represent 3-hydroxy and 4-hydroxy respectively, or
(4) R1, R3 and R4 each represent hydrogen
R2 represents 
xe2x80x83wherein
A represents amino which may be optionally substituted with cycloalkyl, aralkyl, acyl, or aryl which is optionally substituted with one or two substituents selected from the group consisting of halogen, cyano, nitro and amino; or alkyl or aryl each of which may be optionally substituted with halogen,
Y represents CO,
B represents hydrogen,
R5 represents hydroxy, and
R6 and R7 represent 3-hydroxy and 4-hydroxy respectively.
Most preferred compounds include those wherein
(1) R1 represents hydrogen, hydroxy or methoxy,
R2 represents hydrogen, bromine, hydroxy, methyl or amino,
R3 represents hydrogen, hydroxy, methyl, methoxy or amino,
R4 represents hydrogen, halogen or hydroxy,
R5 represents hydroxy, and
R6 and R7 represent 3-hydroxy and 4-hydroxy respectively, or
(2) R1, R3 and R4 each represent hydrogen,
R2 represents 
xe2x80x83wherein
A represents 1,2,3,4-tetrahydroisoquinoline which may be optionally substituted with halogenoalkylcarbonyl or aryl or heteroaryl each of which may be optionally substituted with one or two substituents selected from the group consisting of alkyl, halogenoalkyl, halogen, dialkylamino, phenyl, nitro, amino, isooxazole, pyridine, carboxy, morpholine and methylpiperazine,
Y represents SO2,
B represents hydrogen,
R5 represents hydroxy, and
R6 and R7 represent 3-hydroxy and 4-hydroxy respectively, or
(3) R1, R3 and R4 each represent hydrogen,
R2 represents 
xe2x80x83wherein
A represents amino which is substituted with aryl which is substituted with one or two substituents selected from the group consisting of halogen, cyano, nitro and amino,
Y represents CO,
B represents hydrogen,
R5 represents hydroxy, and
R6 and R7 represent 3-hydroxy and 4-hydroxy respectively.
Typical examples of the flavone derivative of formula (1) according to the present invention are
compound 1) 2-(4-chloro-phenyl)-7-methoxy-chromen-4-one,
compound 2) 7-methoxy-2-phenyl-chromen-4-one,
compound 3) 7-hydroxy-2-phenyl-chromen-4-one.
compound 4) 2-(benzo[1,3]dioxol-5-yl)-3-hydroxy-5-methoxy-chromen-4-one,
compound 5) 2-(benzo[1,3]dioxol-5-yl)-3-hydroxy-6-methoxy-chromen-4-one,
compound 6) 2-(benzo[1,3]dioxol-5-yl)-3-hydroxy-7-methoxy-chromen-4-one,
compound 7) 3-hydroxy-7-methoxy-2-(4-methoxy-phenyl)-chromen-4-one,
compound 8) 2-(4-chloro-phenyl)-3-hydroxy-7-methoxy-chromen-4-one,
compound 9) 6-chloro-3-hydroxy-2-(4-methoxy-phenyl)-8-nitro-chromen-4-one,
compound 10) 6,8-dichloro-3-hydroxy-2-(4-methoxy-phenyl)-chromen-4-one,
compound 11) 3-hydroxy-2-(4-methoxy-phenyl)-chromen-4-one,
compound 12) 6-chloro-2-(3,4-dihydroxyphenyl)-3-hydroxy-4H-chromen-4-one,
compound 13) 6-bromo-2-(3,4-dihydroxyphenyl)-3-hydroxy-4H-chromen-4-one,
compound 14) 2-(3,4-dihydroxyphenyl)-3,6-dihydroxy-4H-chromen-4-one,
compound 15) 6-amino-2-(3,4-dihydroxyphenyl)-3-hydroxy-4H-chromen-4-one,
compound 16) 2-(3,4-dihydroxyphenyl)-3-hydroxy-6-methoxy-4H-chromen-4-one,
compound 17) 6-fluoro-2-(3,4-dihydroxyphenyl)-3-hydroxy-4H-chromen-4-one,
compound 18) 6-methyl-2-(3,4-dihydroxyphenyl)-3-hydroxy-4H-chromen-4-one,
compound 19) 6,8-dichloro-2-(3,4-dihydroxyphenyl)-3-hydroxy-4H-chromen-4-one,
compound 20) 6,8-dibromo-2-(3,4-dihydoxyphenyl)-3-hydroxy-4H-chromen-4-one,
compound 21)6,8-difluoro-2-(3,4-dihydroxyphenyl)-3-hydroxy-4H-chromen-4-one,
compound 22) 6-chloro-2-(3,4-dihydroxyphenyl)-3-hydroxy-7-methyl-4H-chromen4-one,
compound 23) 2-(3,4-dihydroxyphenyl)-3-hydroxy-7-methoxy4H-chromen-4-one,
compound 24) 2-(3,4-dihydroxyphenyl)-3,8-dihydroxy-7-methoxy-4H-chromen-4-one,
compound 25) 2-(3,4-dihydroxyphenyl)-3,8-dihydroxy-4H-chromen-4-one,
compound 26) 2-(3,4-dihydroxyphenyl)-6,7-dimethyl-3-hydroxy-4H-chromen-4-one,
compound 27) 2-(3,4-dihydroxyphenyl)-3-hydroxy-5-methoxy-4H-chromen-4-one,
compound 28) N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]-4-methylbenzenesulfonamide,
compound 29) 4-bromo-N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]benzenesulfonamide,
compound 30) 3-bromo-N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]benzenesulfonamide,
compound 31) N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]-2-naphthalenesulfonamide,
compound 32) N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]-5-(dimethylamino)-1-naphthalenesulfonamide,
compound 33) N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]-2-(1-naphthyl)-1-ethanesulfonamide,
compound 34) 4,5-dibromo-N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-41-1-chromen-6-yl]-2-thiophenesulfonamide,
compound 35) N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]-[1,1xe2x80x2-diphenyl]-4-sulfonamide,
compound 36) N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]-5-(isooxazolyl)-2-thiophenesulfonamide,
compound 37) N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]-5-(2-pyridinyl)-2-thiophenesulfonamide,
compound 38) N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]-3,4-difluorobenzenesulfonamide,
compound 39) N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]-4-(trifluoromethyl)benzenesulfonamide,
compound 40) 4-chloro-N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]-3-nitrobenzenesulfonamide,
compound 41) 3-chloro-N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]-1-propanesulfonamide,
compound 42) N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]-2,4-difluorobenzenesulfonamide,
compound 43) N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]-4-fluorobenzenesulfonamide,
compound 44) N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]-2-(2,2,2-trifluoroacetyl)-1,2,3,4-tetrahydro-7-isoquinolinesulfonamide,
compound 45) 4-({[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]amino}sulfonyl)benzoic acid,
compound 46) N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]-1,2,3,4-tetrahydro-7-isoquinolinesulfonamide,
compound 47) N-cyclohexyl-Nxe2x80x2-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]-sulfide,
compound 48) N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]-4-(4-morpholinyl)-3-nitrobenzenesulfonamide,
compound 49) N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]-4-(4-methyl-1-piperazinyl)-3-nitrobenzenesulfonamide,
compound 50) 3-amino-N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]-4-(4-methyl-1-piperazinyl)benzenesulfonamide,
compound 51) N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]acetamide,
compound 52) N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]benzamide,
compound 53) 4-chloro-N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]benzamide,
compound 54) N-benzyl-Nxe2x80x2-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]urea,
compound 55) N-(4-bromophenyl)-Nxe2x80x2-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]urea,
compound 56) N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]-Nxe2x80x2-phenylurea,
compound 57) N-benzoyl-Nxe2x80x2-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]urea,
compound 58) N-(3-bromophenyl)-Nxe2x80x2-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]urea,
compound 59) N-(2,4-dichlorophenyl)-Nxe2x80x2-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]-urea,
compound 60) N-(3-cyanophenyl)-Nxe2x80x2-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]urea,
compound 61) N-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]-Nxe2x80x2-(4-nitrophenyl)urea,
compound 62) N-(4-aminophenyl)-Nxe2x80x2-[2-(3,4-dihydroxyphenyl)-3-hydroxy-4-oxo-4H-chromen-6-yl]urea,
compound 63) 2-(3,4-dihydroxyphenyl)-3-hydoxy-4H-chromen-4-one,
compound 64) 2-(3,4-dihydroxyphenyl)-3,7-dihydroxy-4H-chromen-4-one,
compound 65) 2-(3,4-dihydroxyphenyl)-3,5,7-trihydroxy-4H-chromen-4-one, and
compound 66) 2-(3,4-dihydroxyphenyl)-3,7,8-trihydroxy-4H-chromen-4-one.
The above compound of formula (1) may also form a pharmaceutically acceptable salt. Such a salt includes non-toxic acid addition salt containing pharmaceutical acceptable anion, for example a salt with inorganic acids such as hydrochloric acid, sulfuric acid, nitric acid, phosphoric acid, hydrobromic acid, hydroiodic acid, etc., a salt with organic carboxylic acids such as tartaric acid, formic acid, citric acid, acetic acid, trichloroacetic acid, trifluoroacetic acid, gluconic acid, benzoic acid, lactic acid, fumaric acid, maleic acid, etc., or a salt with sulfonic acids such as methanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid, naphthalenesulfonic acid, etc. The compound of formula (1) may also exist in the form of hydrate or solvate.
Since the compounds according to the present invention may have asymmetric carbon centers, they can be present in the form of racemate, diastereomer or mixtures thereof. Therefore, the present invention also includes all these isomers and their mixtures.
The compound of the formula (1) according to the present invention may be prepared by process described in the following.
First, the compound of formula (1) wherein R5 is hydrogen, namely the compound of the following formula (1a): 
wherein R1, R2, R3, R4; and R7 are defined as previously described, or a salt thereof may be prepared characterized by
a) cyclizing a compound of the following formula (4): 
wherein R1, R2, R3, R4, R6 and R7 are defined as previously described, in the presence of trifluoroacetic acid to give a compound of the following formula (5): 
wherein R1, R2, R3, R4, R6 and R7 are defined as previously described, then reducing the resulting compound of formula (5) in the presence of a reducing agent in a solvent or
b) cyclizing a compound of the following formula (6): 
wherein R1, R2, R3, R4, R6 and R7 are defined as previously described, in the presence of sodium acetate in a solvent.
The processes are depicted in the following Schemes (1) and (2). 
wherein R1, R2, R3, R4, R6 and R7 are defined as previously described.
In the reaction depicted in Scheme (1), the compound of formula (5) is prepared from cyclization of the compound of formula (4) during which the compound (4) is heated under reflux in the presence of trifluoroacetic acid. This reaction is carried out for an enough time, usually for a day, to complete the reaction depending on the kind of reactant. After the cyclization is completed, the chromane derivative of formula (5) thus obtained is oxidized with a oxidizing agent in a solvent to produce the chromene derivative of above formula (1a). As the solvent, one or more selected from the group consisting of benzene and toluene may be preferably used. As the oxidizing agent, 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ) is used preferably.
In the reaction according to Scheme (2). the compound of formula (1a) wherein R5 is hydrogen is prepared by cyclizing the compound of formula (6) in the presence of sodium acetate in a solvent. At this time, acetic acid is preferably used as the solvent.
Second, the compound of formula (1) wherein R5 is hydroxy, namely the compound of following formula (1b): 
wherein R1, R2, R3, R4, R6 and R7 are defined as previously described, or a salt thereof may be prepared characterized by cyclizing the compound of following formula (4): 
wherein R1, R2, R3, R4, R6 and R7 are defined as previously described, in the presence of a base and hydrogen peroxide in a solvent.
Especially, the compound of formula (1b) wherein R6 and R7 together represent methylenedioxy, namely the compound of formula (1cc): 
wherein R1, R2, R3 and R4 are defined as previously described, can be prepared from the compound of following formula (4a): 
wherein R1, R2, R3 and R4 are defined as previously described, according to the above method.
The process of the above reaction is depicted in the following Scheme (3). 
wherein R1, R2, R3, R4, R6 and R7 are defined as previously described.
In the reaction according to the Scheme (3), the compound of formula (1b) is prepared by cyclizing compound of formula (4) in the presence of hydrogen peroxide. In this process, a solvent and a base are used. As the solvent, the one which does not adversely affect the reaction, such as for example, one or more selected from the group consisting of methanol and ethanol, particularly methanol is used. The base includes one or more selected from the group consisting of sodium hydroxide and potassium hydroxide. Particularly an aqueous sodium hydroxide solution having a concentration of 10% is preferable. This reaction should be conducted inevitably by using hydrogen peroxide which is used in excess amount, preferably in an amount of 5 to 10 times molar equivalents with respect to the compound of formula (4). Suitably, it is used in an aqueous solution of 30%. The reaction may be promoted by optionally adding acid catalyst, such as hydrochloric acid. The reaction is usually carried out at room temperature or under warming, and about 3 hours are usually required to complete the reaction at room temperature.
Deprotection reaction may also be carried out to remove the amino- or hydroxy-protecting group which exists optionally.
That is to say, the compound of formula (1) wherein one or more of the substituents R1, R2, R3 and R4 are hydroxy or amino may be prepared characterized by selectively deprotecting the hydroxy or amino group which is protected by lower alkyl or alkanoyl in the presence of boron tribromide or by hydrolyzing with aqueous sulfuric acid solution or alcohol solvent.
Moreover, the compound of formula (1) wherein R2 is 
namely the compound of the following formula (1h): 
wherein R1, R3, R4, R5 and A, Y, B are defined as previously described, may be prepared by reacting the compound of formula (9): 
wherein R1, R3, R4, R5 and B are defined as previously described, with the compound of formula (10):
Axe2x80x94Yxe2x80x94Xxe2x80x83xe2x80x83(10)
wherein A and Y are defined as previously described and X is a leaving group, or with the compound of formula (11):
Axe2x80x2xe2x80x94Nxe2x95x90Cxe2x95x90Oxe2x80x83xe2x80x83(11)
wherein Axe2x80x2 represents aralkyl, acyl or aryl which is optionally substituted with one or two substituents selected from the group consisting of halogen, cyano and nitro.
Especially, the compound of formula (1h) wherein R5 is hydroxy, namely the compound of formula (1hh): 
wherein R1, R3, R4, A, Y and B are defined as previously described, can be prepared from the compound of formula (9a): 
wherein R1, R3, R1and B are defined as previously described, according to the above method.
The compound of formula (9), especially the compound of formula (9a) used in the above method can be prepared by process described in the following.
A compound of formula (4b): 
wherein R1, R3, R4 and B are defined as previously described, is prepared by reacting 2-hydroxyacetophenone derivative of formula (7a): 
wherein R1, R3, R4 and B are defined as previously described, the piperonal of formula (8a): 
and sodium hydroxide in the aqueous ethanol solution solvent, then thus obtained compound of formula (4b) is reacted with aqueous sodium hydroxide and hydrogen peroxide in methanol solvent to produce a compound of formula (1e): 
wherein R1, R3, R4 and B are defined as previously described.
Then, the above obtained compound of formula (1e) is hydrolyzed with aqueous sulfuric acid solution in alcoholic solvent to give a compound of formula (9a): 
wherein R1, R3, R4 and B are defined as previously described.
The compound of formula (1) wherein one to six of the substituents R1, R2, R3, R4, R6 and R7 are hydroxy may be prepared characterized by selectively deprotecting one to six lower alkoxy groups or lower alkylenedioxy group in the compound of formula (1) in order to convert them into hydroxy of dihydroxy group in the presence of boron tribromide.
For example, the compound of formula (1) wherein R6 and R7 represent 3-hydroxy and 4-hydroxy respectively, namely compound of formula (1d): 
wherein R1, R2, R3, R., and R5 are defined as previously described, may be prepared from the compound of formula (1c): 
wherein R1, R2, R4, R4, and R5 are defined as previously described, according to the reaction as depicted in the following Scheme (4). 
wherein R1, R2, R3, R4, and R5 are defined as previously described.
Especially, the compound formula (1d) wherein R5 is hydroxy, namely the compound of formula (1dd): 
wherein R1, R2, R3 and R4 are defined as previously described, can be prepared from the compound of formula (1cc): 
wherein R1, R2, R3 and R4 are defined as previously described, according to the above method.
In addition, the compound of formula (1cc) wherein R2 is 
namely the compound of formula (1i): 
wherein R1, R3, R4, A, Y and B are defined as previously described, can be prepared from the compound of formula (1hh): 
wherein R1, R3, R4, A, Y and B are defined as previously described, according to the above method.
In the reaction according to Scheme 4, the compound of formula (1d) is prepared by opening the dioxolane ring of the compound of formula (1c) to convert it into dihydroxy group. The opening of dioxolane ring is accomplished by a reaction with boron tribromide which is usually used in an amount of 2 to 5 equivalents, preferably 3 equivalents or more, with respect to the compound of formula (1c). This opening reaction is preferably carried out in a solvent and as the solvent, one or more selected from the group consisting of methylenechloride, dioxane, benzene and toluene, preferably methylenechloride, may be mentioned. The reaction is usually carried out at room temperature or under warming and the reaction is completed at room temperature for about 2 hours.
The compound of formula (4) used as a starting material in the above process for preparation of the compound of formula (1) according to the present invention may be produced by reacting a 2-hydroxyacetophenone derivative of the following formula (7) 
wherein R1, R2, R3, and R4 are defined as previously described, with a compound of the following formula (8): 
wherein R6 and R7 are defined as previously described, in the presence of a base in a solvent.
Especially, the compound of formula (4a) can be prepared by reacting the compound of the formula (7) with piperonal of formula (8a) 
according to the above method.
The above reaction is depicted in the following Scheme (5). 
wherein R1, R2, R3, R4, R6 and R7 are defined as previously described.
In the reaction according to Scheme 5, the compound of formula (4) is prepared by coupling the 2-hydroxyacetophenone derivative of formula (7) with the compound of formula (8) in an amount of 1 to 3 equivalents, preferably 3 equivalents. As the solvent which can be used, one or more selected from the group consisting of water, methanol, ethanol, propanol, butanol and dioxane may be exemplified, and methanol, ethanol or aqueous ethanol solution (80%) is more preferable. As the base, one or more selected from the group consisting of sodium hydroxide, potassium hydroxide, calcium hydroxide, sodium carbonate, potassium carbonate and calcium carbonate, preferably sodium hydroxide, may be mentioned. The amount of base is not particularly restricted, however it is generally used in an amount of 2 to 5 equivalents, preferably 3 equivalents with respect to the compound of formula (7). The reaction is usually carried out at room temperature or under warming and reaction time is variable according to the kind of used solvent or base used, or reaction temperature. The reaction is usually completed at room temperature for about 15 hours.
In the processes as explained in above, the reaction conditions including the amount of reactants, reaction temperature, reaction time, etc. are easily determined depending on the reactants selected by a person having ordinary skill in this art. In general, as for solvent, any one can be used unless it adversely affect the reaction. The reaction temperature may be changeable in variety, however the reaction is preferably carried out at temperatures ranging from 0 to 120xc2x0 C.
The free compound of formula (1) prepared in the reaction according to the present invention may be converted into the above-mentioned salt in a conventional manner well-known in the art. In addition, after the reaction is completed, the product may be isolated or purified by conventional work up procedures such as chromatography, recrystallization, etc.
The compound of the present invention prepared according to the above method has an inhibitory activity against CDK, and thus may be put to a good use as an anti-cancer agent. Accordingly, another object of the present invention is to provide an anti-cancer agent or an agent for treating neurodegenerative disease which comprises the compound of formula (1), pharmaceutically acceptable salt, hydrate, solvate or isomer thereof as an active ingredient together with pharmaceutically acceptable carrier.
In case the compound of the present invention is administered for clinical purpose, it is preferably administered in an amount ranging from 0.01 to 50 mg/kg of body weight a day. The total daily dosage may be administered in one time or over several times. However, the specific dosage for a specific patient can be varied according to the specific compound used, body weight of the subject patient, sex, hygienic condition, diet, time or method of administration, excretion rate, mixing ratio of the medicine, severity of the disease to be treated, etc.
The compound of the present invention may be administered in the form of injections or oral preparations. Injections such as sterilized aqueous or oily suspension for injection may be prepared by using suitable dispersing agent, humectant or suspension agent according to the known method. As solvents to be used for preparing injections, water, Ringer""s fluid and isotonic NaCl solution can be mentioned, and sterilized fixing oil is also used as the solvent or suspension medium. Any non-stimulative fixing oil including mono- or di-glyceride can be used for this purpose, and also fatty acid such as oleic acid can be used for injection formulation.
As the solid preparation for oral administration, capsules, tablets, pills, powders and granules, preferably capsules and tablets can be mentioned. It is desirable for tablets and pills to be formulated into enteric-coated preparation. Solid preparations may be prepared by mixing the active compound of formula (1) according to the present invention with least one carrier selected from a group consisting of inert diluents(e.g. sucrose. lactose, starch, etc.), lubricants(e.g. magnesium stearate), disintegrators, and binders.